Figure 3.
MPRA library preparation and test transfection
(A) Example of non-specific amplification from QIAxcel measurement. In addition to expected ePCR library, non-specific peaks (∼140 bp, ∼220 bp) preclude this sample from further experimental steps.
(B) Successful amplification and library purification of ePCR products (QIAxcel).
(C) Successful library amplification and library purification of tag-seq library from transfected cells (Bioanalyzer).
(D) Agarose gel showing an example of KpnI-digested oligopool after cloning step 2 where cloning step 3 is not required (left lane). Kpn I and XbaI digest to gel-extract fragment for insertion into oligopool from cloning step 1 (right lane).
(E) An example of transfection efficiency (∼>40%) that is required to generate a tag-seq library for a successful and meaningful MPRA. White cells (VSMCs1) depict GFP test transfection.